SFPQ214-598(R542C)/NONO53-312 and the SFPQ276-565/NONO53-312 concentration dependent dimer-tetramer transition

Heidar Koning.

SASDMW8 – The disulphide-linked tetramer of splicing factor, proline- and glutamine-rich and Non-POU domain-containing octamer-binding protein (SFPQ276-598(R542C)/NONO53-312) at 0.78mg/ml

Non-POU domain-containing octamer-binding protein
Splicing factor, proline- and glutamine-rich SFPQ276-598(R542C)/NONO53-312 dimer)
MWexperimental 186 kDa
MWexpected 137 kDa
VPorod 257 nm3
log I(s) 8.02×10-2 8.02×10-3 8.02×10-4 8.02×10-5
Non-POU domain-containing octamer-binding protein Splicing factor, proline- and glutamine-rich SFPQ276-598(R542C)/NONO53-312 dimer) small angle scattering data  s, nm-1
ln I(s)
Non-POU domain-containing octamer-binding protein Splicing factor, proline- and glutamine-rich SFPQ276-598(R542C)/NONO53-312 dimer) Guinier plot ln 8.03×10-2 Rg: 5.2 nm 0 (5.2 nm)-2 s2
(sRg)2I(s)/I(0)
Non-POU domain-containing octamer-binding protein Splicing factor, proline- and glutamine-rich SFPQ276-598(R542C)/NONO53-312 dimer) Kratky plot 1.104 0 √3 sRg
p(r)
Non-POU domain-containing octamer-binding protein Splicing factor, proline- and glutamine-rich SFPQ276-598(R542C)/NONO53-312 dimer) pair distance distribution function Rg: 5.4 nm 0 Dmax: 21 nm

Data validation

Fits and models

log I(s)
 s, nm-1
Non-POU domain-containing octamer-binding protein Splicing factor, proline- and glutamine-rich SFPQ276-598(R542C)/NONO53-312 dimer) DAMMIF model
Synchrotron SAXS data from solutions of SFPQ276-598(R542C)/NONO53-312 at 0.78mg/ml in 20 mM Tris pH 7.5, 250 mM NaCl, were collected on the SAXS/WAXS beam line at the Australian Synchrotron (Melbourne, Australia) using a Pilatus3 S 2M detector at a wavelength of λ = 0.10781 nm (I(s) vs s, where s = 4πsinθ/λ, and 2θ is the scattering angle). One solute concentration of 0.78 mg/ml was measured at 25°C. The data were normalized to the intensity of the transmitted beam and radially averaged; the scattering of the solvent-blank was subtracted.

A tetramer of SFPQ276-598(R542C)/NONO53-312 at 0.78mg/ml which forms when dimers of SFPQ276-598(R542C)/NONO53-312 bind to each other and become cross-linked by the formation of disulfide bond between the coiled-coil domains of SFPQ. The P(r) function and other relevant pieces of data are in agreement with those of a similar construct (SFPQ214-598(R542C)/NONO53-312) resolved by SEC-SAXS with a linear Guinier region (deposited). Additionally, the DAMMIF model of our data overlays reasonably well against a crystal structure of the protein formatted as a tetramer (6WMZ). The rough shape of the molecule appears unchanged as a function of the measured concentration range. We interpret this as the disulfide-linked tetramer persisting over a range of concentrations. Additional data are made available in the full entry zip archive with scattering for the protein at 1.6mg/ml and the corresponding p(r) distribution.

Non-POU domain-containing octamer-binding protein (NONO dimer)
Mol. type   Protein
Organism   Homo sapiens
Olig. state   Dimer
Mon. MW   30.1 kDa
 
UniProt   Q15233 (53-312)
Sequence   FASTA
 
Splicing factor, proline- and glutamine-rich SFPQ276-598(R542C)/NONO53-312 dimer) (SFPQ cysteine-linked)
Mol. type   Protein
Organism   Homo sapiens
Olig. state   Dimer
Mon. MW   38.2 kDa
 
UniProt   P23246 (276-598)
Sequence   FASTA